Methods for treatment of aneurysms

ABSTRACT

Fibrosis, in at least one layer of a vessel wall, can be used to strengthen a vessel wall. Fibrosis can be induced by irradiating a vessel wall with an energy source, or by inducing injury to the vessel wall. In addition to an energy source, photoactivatable agents can also be used such that the energy activates the photoactivatable agent to cause a thickening of the vessel wall. For example, ultra-violet radiation can be used alone or in conjunction with a photoactivatable agent, such as a psoralen compound, to increase the adventitial volume of a blood vessel. Upon exposure to radiation, preferably ultra-violet A radiation, the photoactivatable agent becomes activated and causes compositional and/or structural changes in the adventitia. The invention provides a method of treating aneurysms by thickening the adventitial layer of the vessel wall at the site of the aneurysm.

BACKGROUND OF THE INVENTION

The invention relates to a method for strengthening a vessel wall e.g.,by irradiation with an energy source, or by inducing injury to thevessel wall, such that the injury initiates a cascade of events leadingto fibrosis and a thickening of the vessel wall. In addition to anenergy source, a photoactivable agent can also be used such that theenergy activates the photoactivable agent which causes structuralchanges in the vessel wall. More specifically, the invention relates totreating aneurysms using UV radiation with a psoralen compound.

There are two basic types of blood vessels, arteries and veins, whichcan be distinguished by their structural components. Arteries and veinsboth have several distinct layers which are arranged coaxially. Inarteries, the layers include an inner coat or endothelial layer(intima), an internal elastic lamina, a middle layer (media), and anouter layer (adventitia). Like arteries, veins include an inner layer(intima), a middle layer (media) and an outer layer (adventitia), butthe layers are not as thick as in arteries, and provide less structuralrigidity.

Both arteries and veins are elastic, and are capable of limiteddeformation in response to pressure changes. When a diseased bloodvessel is exposed to hypertension, dilation may occur at variouslocalized regions. Typically these dilatations are produced at theregion in the vessel wall which is weakest, whether inherently or as aresult of disease or trauma. As the dilatation progresses, a morepronounced widening or sac, called an aneurysm, is produced, which mayburst. A large aneurysm can form clots secondary to a reduction in bloodflow in the aneurysmal sac. These clots may emobilize and block distalarteries.

Arteriosclerosis and cystic medial necrosis are two common causes ofaneurysms of the thoracic and abdominal aorta. Thoracic aneurysmscommonly compress and impact surrounding body structures as they expand.They may impact into the lungs, the spinal column, or thegastrointestinal tract.

Berry aneurysms are congenital defects which occur in cerebral arteries,most commonly at the junctions of vessels in the circle of Willis. Theseaneurysms appear to be related to defects in the muscular coat of thevessels. Rupture is common, resulting in intracranial hemorrhage.

Other aneurysms include aortic aneurysms, especially abdominal aorticaneurysms (AAA), which are characterized by transmural aortic walldegeneration leading to dilatation, progressive growth, and eventualrupture.

Treatment of aneurysms typically involves either surgical intervention,such as in thoracic or abdominal aneurysms, or coil ablation, such as inaneurysms of the brain. Excision of the aneurysm, and anastomosis of thevessel may be performed, often using a replacement vessel or anartificial prosthesis. Alternatively, a supporting structure such as astent or other intravascular device may be implanted into the vessel torelieve stress. Examples of stents, include those disclosed in U.S. Pat.No. 4,655,771 issued to Wallsten et al. With the stent positioned at thetreatment site, the stent can be radially expanded into a conformingsurface in contact with a blood vessel wall. The stents may also becovered with a film or a sheath such as, polytetrafluoroethylene (PTFE)as described in U.S. Pat. No. 5,788,626 to Thompson, et al.

Prostheses used to treat aneurysms are also described in, for example,U.S. Pat. No. 4,681,110 issued to Wiktor et al. Wiktor et al. disclosesa flexible tubular liner that is inserted into the aorta to treat aorticaneurysms. The liner has flexible plastic strands designed toelastically expand against the aneurysm and to direct blood flow pastthe aneurysm.

While these methods achieve the general goal of reducing fatality, thesemethods have a drawback in that the aneurysm remains a weak area in theblood vessel wall. Mohr et al. in U.S. Pat. No. 5,921,954 describetreating aneurysms using hardening and softening agents, (e.g.collagen), which are applied at the site of the aneurysm. A radiofrequency energy is then used to harden the agent and cover the weakregion of the blood vessel wall.

Prophylactic methods are used to prevent the formation of aneurysms andrely on reducing the blood pressure, in an effort to reduce mechanicalstress on the vasculature. These methods involve using drugs which canhave undesirable side effects, e.g., cause kidney or liver damage.

Drugs, such as tetracyclines have been used to prevent abnormal vasculardilation, as described in U.S. Pat. No. 5,834,449 issued to Thompson etal. The tetracycline compounds protect the elastic fibers of the mediaby selectively inhibiting the elastolytic activity in this regionthereby preventing its expansion.

While these methods are effective in treating aneurysms, these methodsdo not improve the overall structure of the blood vessel which stillremains weak at the site of the aneurysm and may still be susceptible torupture. Accordingly, one purpose of this invention is to provide amethod of strengthening a blood vessel.

SUMMARY OF THE INVENTION

The invention pertains to methods for strengthening a vessel wall of asubject by thickening or increasing the strength of at least one layerof the blood vessel wall by inducing fibrosis. Strengthening a vesselwall may be accomplished by either applying energy alone, or by applyingenergy with agents, such as compounds and therapeutic agents that inducefibrosis. It has been discovered that certain photoactive compounds,such as psoralen agents play a role in regulating the cell proliferationin the adventitial layer of the blood vessel. This invention is based atleast, in part, on the surprising discovery that blood vessel wallstreated with a psoralen agent, such as 8-methoxypsoralen, (8-MOP)(available commercially as Uvadex, an injectable formulation fromJohnson & Johnson, Exton, Penn.), and subsequently irradiated withlight, e.g., ultra-violet A (UVA) irradiation, will exhibit fibrosis,especially in the adventitial layer.

The vessel wall can be strengthened by altering the cellular andmolecular processes to thicken one or more layers of the vessel wall,for example, by inducing fibrosis in the adventitia of a blood vessel,thereby enabling it to return to a more normal, less weakened state. Themethod of the invention can be used to treat aneurysms by inducing thefibrosis in the adventitial layer of a blood vessel.

Accordingly, in one aspect, the invention pertains to a method forstrengthening the vessel wall of a subject by identifying a region ofweakness in a vessel wall, the region of weakness comprising at leastone target layer; and applying energy to the region of weakness in anamount effective to induce fibrosis in a target layer, to therebystrengthen the vessel wall. The region of weakness can be identifiedusing techniques such as ultrasound analysis, X-ray analysis,computerized tomography, magnetic resonance imaging (MRI), orangiography.

Energy can be applied to the region of weakness by irradiating theregion of weakness with X-ray irradiation in an amount effective toinduce fibrosis in a target layer.

Other forms of energy applied in an amount effective to induce fibrosisin a target layer also include, but not limited to, UV irradiation, IRirradiation, microwave irradiation, heat irradiation and RF irradiation.

The method can further comprise, administering a therapeuticallyeffective amount of an agent to a subject, such that the agent is takenup by at least one target layer of the vessel wall. In one embodiment,the agent can be a photoactivatable agent, such that thephotoactivatable agent is activated upon irradiation to induce fibrosisin a target layer.

In another aspect, the invention pertains to a method for strengtheninga vessel wall of a subject by administering a therapeutically effectiveamount of a photoactivatable agent to a subject, such that the agent istaken up by at least one layer of the vessel wall, and irradiating atarget region of the vessel wall, such that the photoactivatable agentis activated to strengthen the vessel wall.

The photoactivatable agent can be administered using any known methodsfor administrating therapeutic agents, for example, systemic, local,oral administration, and the like. In one embodiment, the administeringstep comprises systemically administering the photoactivatable agent. Inanother embodiment, the administering step comprises locallyadministering the photoactivatable agent.

The photoactivatable agent can be any agent that is activated by lightenergy. In the activated state, the photoactivatable agent is capable ofcausing changes in the cellular and molecular processes in the localizedmicroenvironment, e.g., structural changes by altering cellproliferation in at least one layer of the vessel resulting in a changeof thickness in the layer. In one embodiment, the photoactivatable agentis a psoralen agent or a derivative thereof.

The photoactivatable agent can be activated using a light source in avariety of ways. Photoactivation can occur by irradiating a targetregion internally by a light source applied at the target regionin-vivo. Photoactivation may also occur by irradiating the target regionusing an external light source. The entire area of the subject can beirradiated externally or, a desired localized area can be irradiatedexternally. In one embodiment, the irradiating step comprisesirradiating the target region internally using a light deliverycatheter. In another embodiment, the irradiating step comprisesirradiating the target region using a light delivery catheter withoutoccluding fluid flow. In yet another embodiment, the irradiating stepcomprises irradiating the target region externally using an externallight delivery source. In a preferred embodiment, the irradiating stepcomprises irradiating the target region with UV light, preferably withlight having a wavelength of about 240 to 370 nanometers. In anotherembodiment, the irradiating step comprises irradiating the target regionto increase the area of the vessel wall outer layer, e.g., theadventitia.

In another aspect, the invention pertains to a method for increasing theadventitial mass of a blood vessel wall within a target region byadministering a therapeutically effective amount of a photoactivatableagent to a subject, such that the agent is taken up by the adventitiallayer, and irradiating a target region of the blood vessel wall so thatthe photoactivatable agent is activated to increase the adventitialvolume.

In another aspect, the invention pertains to a method for treating ananeurysm by increasing the adventitial volume of a blood vessel byadministering a therapeutically effective amount of a photoactivatableagent to a subject, such that the agent is taken up by the adventitialregion of the blood vessel, and irradiating the site of the aneurysm sothat the photoactivatable agent increases the adventitial volume.

Strengthening a vessel wall of a subject by increasing the volume of atleast one layer of the blood vessel wall may also be attained byirradiating the target region with light energy of a specificwavelength. The irradiation alone can be sufficient to activate cellularand molecular process that result in an increase in a vessel wall layer.

Accordingly, in another aspect, the invention pertains to a method forstrengthening a vessel wall of a subject by irradiating a target regionwith UVC irradiation, so that the UVC irradiation induces a structuralchange in at least one layer of the vessel wall. Additionally, theinvention pertains to a method for increasing the adventitial volume ofa blood vessel wall by irradiating the target region with UVCirradiation, and more specifically, to a method for treating an aneurysmby increasing the adventitial volume of a blood vessel by irradiatingthe site of the aneurysm with UVC irradiation.

BRIEF DESCRIPTION OF THE DRAWINGS

FIG. 1 is a graph showing the effect of Psoralen-Ultra Violet A (PUVA)therapy on the adventitial area of a blood vessel.

DETAILED DESCRIPTION

The present invention pertains to methods for strengthening a vesselwall of a subject by identifying a region of possible or actual weaknessin the vessel wall, the region of weakness comprising at least onetarget layer; and applying energy to region of weakness in an amounteffective to induce fibrosis in a target layer, to thereby strengthenthe vessel wall.

The present invention also pertains to methods for strengthening avessel wall of a subject by administering a therapeutically effectiveamount of a photoactivatable agent to the subject, such that the agentis taken up by at least one layer of the vessel wall, and irradiating atarget region of the vessel wall, such that the photoactivatable agentis activated to strengthen the vessel wall. The data described hereindemonstrates that a psoralen compound (e.g., 8-MOP, available as Uvadex,from Johnson and Johnson) and UVA irradiation causes proliferation ofthe adventitial layer of a blood vessel. These results are unexpectedsince psoralen compounds in general have little or no effect onproliferation of cells in the intima or adventitia of a blood vessel.

So that the invention may more readily be understood, certain terms arefirst defined.

The term “strengthening” as used herein refers to restoring the vesselwall to a more normal, less weakened state in a subject. Strengtheningmay occur by altering cellular and molecular processes to thicken one ormore layers in the vessel wall. Induction of fibrosis may lead tostrengthening of a vessel wall. For example, a vessel with an aneurysmis detrimental to the subject because the vessel wall comprising theaneurysm becomes thin and weak, presenting the risk of rupture ordissection. Strengthening the vessel wall involves increasing the areaof the vessel wall surrounding the aneurysm so that it becomes thickerand less susceptible to rupture.

The phrase “region of weakness” as used herein refers to an area in avessel wall that requires strengthening, including, for example, bloodvessel regions that are dilated or otherwise deviated in size or shapefrom a normal or reference value as well as regions that are identifiedby MRI scanning or other imaging modality.

The phrase “target layer” as used herein refers to at least one layer ofa vessel wall in which cellular and molecular processes, such asfibrosis, occur.

The term “energy” as used herein refers to any source from theelectromagnetic spectrum that is applied for a duration, and anintensity to cause the desired result. Examples of different forms ofenergy include but are not limited to X-rays, microwaves, UV radiation,IR radiation, visible light, and radio frequency waves. Energy can alsobe applied by extracting heat, e.g., with a cryogenic catheter. Thephrase “amount effective to induce” as used herein refers to theduration and dosage of energy sufficient to initiate a cascade of eventsleading to fibrosis.

The term “fibrosis” as used herein refers to the art recognized meaningof the term. Fibrosis is a complex process involving different celltypes such as fibroblasts, myofibroblasts, macrophages. Fibrosis is aresponse to injury in which new extracellular matrix is laid downproducing dense amounts of collagen required for wound healing. Thecascade of events in wound healing involves activation or release ofmolecules such as cytokines, growth factors and adhesion molecules. Theextracellular matrix growth factors TGFβ, platelet derived growth factor(PDGF), and basic fibroblast growth factor (bFGF) appear to initiate andsustain fibrosis. In particular, TGFβ stimulates collagen andfibronectin formation, suppresses collagenase and induces production ofcollagenase inhibitors. Wound healing occurs immediately after injuryand provides a means to remove the damaged tissue from the wound. Soonafter, fibroblasts from the surrounding tissue move into the area oftissue injury leading to an increase in fibroblast and cellularity atthe site of injury. The fibroblasts proliferate in the injured area andactively produce macromolecular compounds such as collagen andproteoglycans which are secreted into the extracellular matrix of thetarget layer. The newly synthesized collagen fibrils are cross-linked bylysyl oxidase and provide structural integrity to the wound. At a finalstage, the remodeling stage, the previous randomly organized collagenfibrils are aligned in the direction of mechanical tension and becomemore organized to increase the mechanical strength of the wounded area.Fibrosis can be induced by physical (e.g., heat, cold) and/or chemical(e.g., sodium hydroxide, hydrogen peroxide) stimuli.

The phrase “therapeutically effective amount” as used herein refers toan amount effective, at dosages and for periods of time necessary, toachieve the desired result. A therapeutically effective amount of thephotoactivatable agent or derivatives thereof may vary according tofactors such as the disease state, age, and weight of the subject, andthe ability of the photoactivatable agent or derivatives thereof (aloneor in combination with one or more other agents) to elicit a desiredresponse in the subject. Dosage regimens may be adjusted to provide theoptimum therapeutic response. A therapeutically effective amount is alsoone in which any toxic or detrimental effects of the photoactivatableagent or derivatives thereof are outweighed by the therapeuticallybeneficial effects.

The phrase “photoactivatable agent” as used herein refers to a materialwhich becomes activated by light energy. In the activated state, thephotoactivatable agent is capable of causing changes in the cellular andmolecular processes in the localized microenvironment, e.g., structuralchanges in the tissue layer, such as, altering cell proliferation in theadventitial layer that results in an increase of area in this layer. Inone embodiment, the photoactivatable agent is a chromophore. Suitablechromophores are generally selected for absorption of light that isdeliverable from common radiation sources (e.g. UV light ranging from240-370 nm). Examples of chromophores which are photoresponsive to suchwavelengths include, but are not limited to, acridines, nitroaromaticsand arylsulfonamides. One preferred class of chromophores comprisespsoralen compounds.

The phrase “target region” as used herein refers to an area of thevessel wall in need of treatment. When the target area is irradiated byan energy source, it undergoes a compositional or structural change thatcorrects the defect. The target region is capable of undergoing suchchanges when irradiated with an energy source alone or in combinationwith a photoactivatable agent.

Various aspects of the invention are described in more detail in thefollowing subsections:

I. Identification of Regions of Weakness

Standard procedures for identifying regions of weakness in a vessel wallcan be employed. For example, visualization using CT scans, X-rayanalysis, ultrasound analysis, Magnetic Resonance Imaging (MRI), andcomputed tomography scans. Having identified the location of the regionof weakness, appropriate procedures can be undertaken to treat theregion of weakness by inducing fibrosis. Appropriate procedures caninclude, for example, inserting a catheter at the region of weakness(e.g., an aneurysm), and transmitting a source of energy that inducesfibrosis.

The method of the invention also permits early intervention andtreatment of regions of weakness. Aneurysms, for example, can be treatedby strengthening the vessel wall as soon as the aneurysm has beenidentified. Typically, the recommended procedure for treating aneurysmsinvolves surgery or applications of stents. Before these procedures areundertaken, the patient is usually monitored until the aneurysm is of asize suitable for treatment. By strengthening the vessel wall as soon asthe aneurysm is identified, the risk associated with rupture, embolismsor dissection of the aneurysm is reduced.

II. Injury Induction

Application of a stimuli that results in tissue injury produces acascade of events that promotes repair of the damaged tissue. Atherapeutic method has been developed in which the response to injury isused to strengthen a vessel wall (e.g., blood vessel wall). The cascadeof events involves activation and release of molecules such ascytokines, growth factors and adhesion molecules. In particular,fibrosis is a response to tissue injury in which new extracellularmedium is laid down producing dense amounts of collagen. Stimuli thatcauses injury can be used to induce activation of the cascade. Thestimuli may be from any range of the electromagnetic spectrum (e.g.,heat, light, sound, microwaves, radio frequency waves, or irradiation,(X-ray, UV, IR)) which can be applied using standard means. Stimuli thatcauses injury can also be chemical such as hydrogen peroxide, sodiumhydroxide, and the like, or pressure related, for example, pressure on avessel wall due to procedures like angioplasty.

In one embodiment, exposure of the vessel wall to heat results in injuryto the vessel wall. Heat can be applied over a temperature range ofabout 40° C. to about 60° C., preferably about 40° C. to about 50° C.,and most preferably about 42° C. to about 45° C. for a period of timethat results in injury (e.g., 1 minute, 30 seconds, 2 minutes) to avessel wall. Heat can be applied by direct contact of the vessel wallwith the heat source, for example, a device such as a cathetercontaining a thermocouple. In some cases localized heating may beachieved using a laser, infra-red, or microwave irradiation. The heatingconditions should be sufficient to elevate the temperature of thelocalized area to at least about 42° C. for a period of time thatresults in injury without killing the cells in the vessel wall.

In another embodiment, cryotherapy techniques may also be used to reducethe temperature of the vessel wall to induce injury. Methods forreducing the temperature of a vessel wall are known in the art (Seee.g., U.S. Pat. No. 5,902,299.) The temperature of the vessel can bereduced to cause injury using a catheter that delivers a medium, such asgas, fluid or a mixture thereof at a very low temperature between 14° C.and about −10° C. to the vessel wall.

In another embodiment, exposure of the vessel wall to chemicals cancause injury. Chemical inducers, such as hydrogen peroxide, (Canrgeon etal. (1997) Exp. Cell. Res., 20: 30-37) transition metals such as copper,zinc, cadmium (See e.g., Levinson et al. (1980) Biochem. Phys. Acta.,606: 170-180) can be applied to the vessel wall at concentrationssufficient to cause injury without killing the cells.

In addition to treating arteries, the method of the invention can alsobe used for other applications for example, treating veins, ureters,urethra, bronchi, biliary, pancreatic duct systems, the gut, eustatian,spermatic and fallopian tubes. The method of the invention may be usedto treat an lesion in hollow vessels composed of several tissue layers.

The method of the invention can also be used prophylactically, toprevent the growth of aneurysms, or to induce a reduction in the size ofthe aneurysm. In, the embodiments, the method of the invention can alsobe used to inhibit or reduce inflammation in the vessel.

III. Agents

The present invention can be practiced in conjunction with theadministration of photoactivatable agents that enhance the effects ofirradiation. Psoralens are tricyclic compounds formed by the linearfusion of a furan ring with a coumarin. Psoralens can intercalatebetween the base pairs of double-stranded nucleic acids, formingcovalent adducts to pyrimidine bases upon absorption of longwaveultraviolet light. (See e.g., Cimino et al. (1985) Ann. Rev. Biochem.54:1151-1193 and Hearst et al. (1984) Quart. Rev. Biophys. 17:1-44).Psoralens are a group of compounds that exhibit photoactivation. (Seee.g., Parrish et al. (1974) N. Engl. J. Med, 291: 1207-1211 and Edelsonet al. (1987) N. Engl. J. Med., 316: 297-303), herein incorporated byreference, for detailed descriptions of psoralen compounds. Psoralensand furocoumarins (furanes fused to coumarin and derivatives thereof)are a preferred class of photoactivatable agents. A psoralen compoundcan be administered to the subject prior to irradiating the targetregion. The psoralen compound will be preferentially absorbed by thelayers of the vessel wall, thus rendering them more susceptible to theUV light.

The interactions of psoralen compounds with DNA have previously beendescribed (See e.g., Malane, et al. (1991) Ann. N.Y. Acad. Sci.636:196-208 also incorporated herein by reference). If a psoralencompound is administered orally, the psoralen compound is absorbed fromthe digestive tract, reaching peak levels in the blood and other tissuesin one to four hours. Psoralen compounds are excreted within 24 hoursfollowing oral administration. The psoralen compound is inert prior toexposure to ultraviolet or visible light irradiation and is transientlyactivated into an excited state following irradiation. The transientlyactivated compound is capable of photomodifying biological molecules(e.g., DNA, protein) and generating other reactive species, (e.g.,singlet oxygen), which are capable of modifying other cellularcomponents.

One type of radiation useful for photoactivation of psoralen compoundsis ultraviolet A irradiation. Alternatively, visible light having awavelength greater than about 420 nm can be useful with certain psoralencompounds (See Gasparro, et al., (1993), 57:1007-1010 hereinincorporated by reference).

Without being limited to any particular mechanism or explanation, it isbelieved that the DNA cross-linking by psoralen compounds proceeds by atwo step process. Following administration of the psoralen compound intothe subject, the psoralen compound first intercalates within the doublehelix of intracellular DNA or RNA. Following intercalation, the psoralencompound is covalently added to the polynucleic acid by light energywithin the absorption band of the psoralen compound. Either psoralen-RNAor psoralen-DNA monoadducts or cross-links are created upon illuminationof the intercalated species. By forming covalent cross-links withbase-pair structures, psoralen compounds alter the metabolic activity ofa cell (See e.g., Cimino, et al. (1985), Ann. Rev. Biochem., 54:1154-93also herein incorporated by reference).

Examples of photoactivatable agents useful in the present invention caninclude, but are not limited to, 5-methoxypsoralen (5-MOP),8-methoxypsoralen (8-MOP), 4,5′,8-trimethylpsoralen (TMP),4′-aminomethyl-4,5′,8-trimethylpsoralen (AMT),5-chloromethyl-8-methoxypsoralen (HMT), angelicin (isopsoralen),5-methylangelicin (5-MIP), 3-carboxypsoralen, porphyrin,haematoporphyrin derivative (HPD), photofrin II, benzoporphyrinderivative (BPD), protoporphyrin IX (PpIX), dye haematoporphyrin ether(DHE), polyhaematoporphyrin esters (PHE),13,17-N,N,N-dimethylethylethanolamine ester of protoporphyrin (PH1008),tetra(3-hydroxyphenyl)-porphyrin (3-THPP), tetraphenylporphyrinmonosulfonate (TPPS1), tetraphenylporphyrin disulfonate (TPPS2a),dihaematoporphyrin ether, mesotetraphenylporphyrin,mesotetra(4N-methylpyridyl)prophyrin (T4MpyP),octa-(4-tert-butylphenyl)tetrapyrazinoporphyrazine (OPTP),phthalocyanine, tetra-(4-tert-butyl)phthalocyanine (t₄-PcH₂),tetra-(4-tert-butyl)phthalocyanatomagnesium (t₄-PcMg), chloroaluminumsulfonated phthalocyanine (CASPc), chloroaluminum phthalocyaninetetrasulfate (A1PcTS), mono-sulfonated aluminum phthalocyanine (A1SPc),di-sulfonated aluminum phthalocyanine (A1S2Pc), tri-sulfonated aluminumphthalocyanine (A1S3Pc), tetra-sulfonated aluminum phthalocyanine(A1S4Pc), silicon phthalocyanine (SiPc IV), zinc II phthalocyanine(ZnPc), pyrene, bis(di-isobutyl octadecylsiloxy)silicon2,3-naphthalocyanine (isoBOSINC), germanium IV octabutoxyphthalocyanine(GePc), rhodamine 101 (Rh-101), rhodamine 110 (Rh-110), rhodamine 123(Rh-123), rhodamine 19 (Rh-19), rhodamine 560 (Rh-560), rhodamine 575(Rh-575), rhodamine 590 (Rh-590), rhodamine 610 (Rh-610), rhodamine 640(Rh-640), rhodamine 6G (Rh-6G), rhodamine 700 (Rh-700), rhodamine 800(Rh-800), rhodamine B (Rh-B), sulforhodamine 101, sulforhodamine 640,sulforhodamine B, coumarin 1, coumarin 2, coumarin 4, coumarin 6,coumarin 6H, coumarin 7, coumarin 30, coumarin 47, coumarin 102,coumarin 106, coumarin 120, coumarin 151, coumarin 152, coumarin 152A,coumarin 153, coumarin 311, coumarin 307, coumarin 314, coumarin 334,coumarin 337, coumarin 343, coumarin 440, coumarin 450, coumarin 456,coumarin 460, coumarin 461, coumarin 466, coumarin 478, coumarin 480,coumarin 481, coumarin 485, coumarin 490, coumarin 500, coumarin 503,coumarin 504, coumarin 510, coumarin 515, coumarin 519, coumarin 521,coumarin 522, coumarin 523, coumarin 535, coumarin 540, coumarin 540A,coumarin 548, 5-ethylamino-9-diethylaminobenzo α-phenoxazimium (EtNBA),5-ethyl-amino-9-di ethyl-aminobenzo α-phenoxazinium (EtNBS),5-ethylamino-9-diethylaminobenzo α-pheno-selenazinium (EtNBSe),chlorpromazine, chlorpormazine derivatives, chlorophyll derivatives,bacteriochlorophyll derivatives, metal-ligand complexes,tris(2,2′-bipyridine)ruthenium (II) dichloride (RuBPY),tris(2,2′-bipyridine)rhodium (II) dichloride (RhBPY),tris(2,2′-bipyridine)platinum (II) dichloride (PtBPY), pheophorbide,merocyanine 540, vitamin D, 5-amino-laevulinic acid, photosan, chlorine6, chlorin e6 ethylene-diamide, mono-L-aspartyl chlorin e6, andphenoxazine Nile blue derivatives, stilbene, stilbene derivatives, and4-(N-2(2-hydroxyethyl)-N-methyl)-aminophenyl)4′-(6-hydroxyhexylsulfonyl)-stilbene(APSS). In a preferred embodiment, the photoactivatable agent is apsoralen.

IV. Administration of Agents

Suitable agents such as photoactivatable agents, may be administeredusing any known method. Administration may involve needle injectionsinto cells, tissues and fluid spaces or blood vessels. Examples ofadministering photoactivatable agents include but are not limited tointravenous, intramuscular, aerosol, oral, topical, systemic, ocular,intraperitoneal and/or intrathecal.

In one embodiment, the photoactivatable agent can be administeredsystemically. Systemically administered photoactivatable agents e.g.psoralen compounds, penetrate the nuclear membrane of cells and canintercalate with the nuclear DNA in target tissue cells. Followingintercalation with the target tissue's nuclear DNA, the psoralencompound is photoactivated with ultraviolet light or short wavelengthvisible light. (See, e.g., Gasparro, et al. (1993), Photochem.Photobiol. 57:1007-1010.)

In another embodiment, the photoactivatable agent can be administeredlocally. Typically, local administration of the agent is achievedthrough conventional devices such as catheters, laparoscopes,endoscopes, cannulae, direct injection. Approaches for local,intravascular, site-specific administration of agents have also includeddirect deposition of such agents into the vessel wall through anintravascular delivery system. These intravascular delivery systemsgenerally employ balloon catheters which are easily guided through bloodvessels to a region in need of treatment and can then be inflated tofully contact and dilate the entire surrounding vessel wall. Atherapeutic agent can then be delivered to the surrounding vessel wall,for example, by diffusion through the balloon or by hydrostaticpressure, as occurs when using a porous balloon catheter.

Other balloon catheters which have been used for drug delivery to bloodvessel walls are drug-coated catheters (e.g., hydrogel catheters). Uponinflation of the balloon in a blood vessel, the therapeutic agent is“pressed” onto or into the surrounding vessel wall. (See, e.g., Sheriff,et al., (1993), J. Am. Coll. Cardiol. 21:188A herein incorporated byreference). Typically, the balloon must be chaperoned by a protectivesheath as the catheter is advanced toward the target vessel.

In a preferred embodiment, a non-occluding catheter can be used.Non-occluding catheters are described in a co-pending application byKasinkas et al. entitled “Non-Occluding Light Delivery Catheter” and isincorporated herein by reference. The non-occluding catheter is designedto deliver the agent to the target region in the vessel wall whilepermitting blood flow through the vessel wall.

V. Activation of Photoactivatable Agents

Photoactivatable agents can be activated using a monochromatic lightsource such as a laser. In one embodiment, the photoactivatable agent isactivated internally. For example, the light output may be coupled to aninvasive, light delivery catheter for conduction and delivery to aremote target region. Such interventional light delivery catheters arewell known in the art and are described, for example, in U.S. Pat. No.5,169,395 issued to Narisco, et al., U.S. Pat. No. 5,196,005 issued toDoiron, et al., U.S. Pat. No. 4,773,999 issued to Spear, et al. and U.S.Pat. No. 5,231,684 issued to Narisco, et al. Other devices which arefrequently used in conjunction with a light source and light deliverycatheter include drug delivery devices and/or a balloon perfusioncatheter (See e.g., U.S. Pat. No. 5,213,576 issued to Abiuso, et al.)

In a preferred embodiment, a non-occluding catheter can be used.Non-occluding catheters are described in a co-pending application byKasinkas et al. entitled “Non-Occluding Light Delivery Catheter” and isincorporated herein by reference. The non-occluding catheter can be usedto deliver light or UVA energy to the target region in the vessel wallwhile permitting blood flow through the vessel wall.

In another embodiment, the photoactivatable agent is activatedexternally. For example, using a monochromatic light source applied toan area of the subject's body requiring treatment, for example, thechest area. Such external light sources are well known in the art, forexample, a monochromatic light source, e.g. a UV lamp.

Activation of photoactivatable agents may also be achieved by injectinga form of liquid light into the vascular system of the subject. Examplesof light-emitting liquids are the bioluminescent system of fireflylucerin/lucerase and the chemiluminescent system of the CyalumeLightstick manufactured by the American Cyanamid Company. Luciferin andluciferase are water soluble, and light is emitted when adenosinetriphosphate, which is water soluble, is added to these substances. Abuffer such as glycine and the metal ion, magnesium, is usually presentin the solution to facilitate the reaction. Intravenous injection ofthese materials, obtained commercially from Sigma Chemical Company, intodogs has produced no deleterious side effects.

Aqueous peroxyoxylate chemiluminescent liquids are another example oflight emitting liquids and may be injected into the bloodstream of ratsand rabbits without producing any side effects. The liquid reactantstypically include a triflyl oxamide and hydrogen peroxide along withsulfonated rubrene as a fluorescer and Deceresol NI as a surfactant.

A potential advantage of the use of liquid light is that all diseasedvessels can be perfused with the liquid light, with one intravascularinjection of the liquid light. Knowledge of the exact location of targetregion is unnecessary, since all would be exposed to the light.

One skilled in the art will appreciate further features and advantagesof the invention based on the above-described embodiments. Accordingly,the invention is not to be limited by what has been particularly shownand described, except as indicated by the appended claims. Allpublications and references cited herein are expressly incorporatedherein by reference in their entirety.

Example

The Effect of PUVA on Blood Vessel Walls

To evaluate the effect of PUVA therapy (psoralen photoactivated with UVAlight) on blood vessel walls, 17 healthy, juvenile pigs receivedintracoronary PUVA therapy immediately following balloon-injury. A totalof 33 coronary arteries were treated and evaluated histologicallytwenty-eight days later.

The animals, ranging in weight from 29.8 kg to 45.4 kg, wereanesthetized and prepared for treatment according to standard laboratoryprocedures. Quantitative coronary angiography was performed to identifysuitable arterial segments for treatment and to provide baselineinformation. Arteries selected for study were injured using a standardcoronary balloon catheter inflated three times to 120% nominal vesselsize.

All animals received Uvadex (Johnson & Johnson, Exton, Penn.), anintravenous formulation of 8-methoxypsoralen, which was administered ata constant infusion rate to yield the specified dose at sixty minutesfollowing the start of infusion. This constant infusion was maintaineduntil light delivery was completed.

At approximately sixty minutes following the start of Uvadex infusion,the intravascular light delivery procedure was started. The lightdelivery system consisted of a Compass, frequency tripled YAG laser,(Coherent, Santa Clara, Calif.) optically coupled to a light guide anddelivery catheter specifically designed to deliver light to coronaryarteries (Illumenex Corporation, Plymouth, Minn.). The light guide,delivery catheter, and light delivery procedure are described in moredetail in U.S. Pat. Nos. 5,620,438 and 5,833,682 both issued to Amplatzet al.

As shown in Table 1 below, six PUVA doses (including controls) wereevaluated by varying the amount of psoralen administered to the animaland the amount of light (355 nm, 40 kHz pulsed) delivered to the artery.Although light treatment parameters vary with vessel diameter, thedelivery of a 10 J/cm² dose to a 3.2 mm diameter artery typicallyinvolved 170 mW delivered for 210 seconds. Other light doses wereachieved by varying the amount of power and/or light exposure time.Arteries assigned to the control received a constant infusion of Uvadex,and underwent an equivalent light delivery procedure, except that thepower level was set to 0 mW.

TABLE 1 PUVA Doses Psoralen UVA Number of Arteries  1.0 mg/kg  10 J/cm²5  0.5 mg/kg  10 J/cm² 5 0.125 mg/kg  10 J/cm² 9 0.250 mg/kg   1 J/cm² 20.125 mg/kg 3.3 J/cm² 2  1.0 mg/kg   0 J/cm² 3  0.5 mg/kg   0 J/cm² 30.125 mg/kg   0 J/cm² 4

To determine the concentration of Uvadex in plasma, blood samples weretaken at fifteen-minute intervals during and following the completion ofthe intravenous infusion. Additional blood samples were taken duringlight delivery for purposes of determining the PUVA treatment dose.Blood samples were processed and blind-labeled for analysis to determinepsoralen plasma concentrations (Jefferson University Photobiology Lab,Philadelphia, Penn.) according to methods described by Gasparro et al.,(1988) J. Invest. Dermatol. 90: 234-236.

At approximately 28 days after treatment, animals were euthanizedaccording to standard laboratory procedures immediately followingcompletion of quantitative coronary angiography. The hearts from eachanimal were perfusion fixed with formalin and the coronary arteriesexcised. Histological sections were prepared from the excised arteries,and morphometric data was obtained from these sections according tostandard procedures.

To evaluate the effect of PUVA therapy to strengthen arterial walls, thearea comprising the adventitial layer of the artery (Adventitial Area)was calculated for each of the 189 histological sections prepared fromthe 33 arteries enrolled in the study. An increase in adventitial areais strongly indicative of strengthening of the vessel wall.

Analysis I

To evaluate whether PUVA therapy effected the Adventitial Area, each ofthe 189 arterial sections was assigned to a Treatment or Control Group,and differences between the groups was assessed. As shown in Table 2below, the average Adventitial Area for 155 arterial sections of theTreatment Group was 5.6 mm^(2,) whereas the 34 sections of the ControlGroup averaged only 2.7 mm². An analysis of the difference between thesample means (one-tail, 5%, assuming unequal variances) indicated thatthe difference, approximately 2.9 mm^(2,) was statistically significant,with a probability value (p-value) of 4.1×10⁻¹⁵.

TABLE 2 Adventitial Area, Treatment v. Controls Treatment Group ControlGroup Number of Arteries 23 10 Number of Sections 155 34 Average 5.6 2.7Variance 10.9 1.4 Sample Mean Difference 2.9 Standard Error 0.33229 tStatistic 8.749 P-value 4.1 × 10⁻¹⁵

Analysis 2

To evaluate whether the effect of PUVA therapy on Adventitial Area wasdose-dependent, each of the 189 arterial sections was assigned to one offour Treatment Dose Groups according to criteria listed in Table 3below. For purposes of analysis, a Treatment Dose was defined to be theproduct of the psoralen concentration in plasma (based on blood samplestaken at the time of light delivery) and energy density (based on powermeasurements taken from the delivery catheter following light delivery),and is given in units of nanograms/ml×J/cm².

TABLE 3 Treatment Dose Groups Treatment Dose Group Grouping CriteriaHigh 5,000 < Treatment Dose Medium 1,000 < Treatment Dose <= 5,000 Low   0 < Treatment Dose <= 1,000 Control Treatment Dose = 0

The results from the study are summarized in Table 4 below. Theseresults demonstrated that the average Adventitial Area increased withTreatment Dose Group, and that the differences were statisticallydifferent between all paired groups except Low and Controls.

TABLE 4 Adventitial Area by Treatment Dose Group High Medium Low ControlNumber of Arteries 6 7 10 10 Number of Sections 44 55 56 34 Average 8.45.9 3.3 2.7 Variance 8.9 9.4 1.9 1.4 Sample Mean Difference 2.50 2.740.45 Standard Error 0.61055 0.45098 0.27088 t Statistic 4.098 6.0761.649 P-value 4.4 × 10⁻⁵ 2.4 × 10⁻⁸ 0.052

Analysis 3

To further evaluate whether the effect of PUVA therapy on AdventitialArea was dose-dependent, Adventitial Area was plotted versus TreatmentDose for the 189 arterial sections, as illustrated in FIG. 1. A simplelinear regression of this data indicated that there was a positive,statistically-significant relationship between the Adventitial Area andTreatment Dose. The statistical analysis data corresponding to FIG. 1 ispresented in Tables 5 A-C, below.

TABLE 5A Statistical Analysis Data Regression Statistics Multiple R68.2% R Square 46.5% Adjusted R Square 46.2% Standard Error 2.37Observations 189

TABLE 5B Statistical Analysis Data ANOVA df SS MS F Significance FRegression 1 909 909 162 3.4 × 10⁻²⁷ Residual 187 1046 6 Total 188 1956

TABLE 5C Statistical Analysis Data Regression Standard ParametersCoefficients Error t Statistic P-value Intercept 3.2 0.229 13.9 1.1 ×10⁻³⁰ Treatment Dose 7.1 × 10⁻⁴ 5.6 × 10⁻⁵ 12.7 3.5 × 10⁻²⁷

Collectively, the results of this study demonstrated that (i)intracoronary PUVA therapy increased the size of the adventitial layerof the artery, and (ii) such adventitial growth was dose dependent.

What is claimed is:
 1. A method for strengthening a vessel wall in anarea of an aneurysm of a subject comprising: identifying a region ofweakness in a vessel wall of an aneurysm, the region of weaknesscomprising at least one target layer; applying an agent in combinationwith energy to the region of weakness; and inducing fibrosis in a targetlayer, to thereby strengthen the vessel wall.
 2. The method of claim 1,wherein the step of identifying the region of weakness comprises usingultrasound analysis.
 3. The method of claim 1, wherein the step ofidentifying the region of weakness comprises using X-ray analysis. 4.The method of claim 1, wherein the step of applying energy to the regionof weakness comprises irradiating the region of weakness with X-rayirradiation in an amount effective to induce fibrosis in a target layer.5. The method of claim 1, wherein the step of applying energy to theregion of weakness comprises irradiating the region of weakness with UVirradiation in an amount effective to induce fibrosis in a target layer.6. The method of claim 1, wherein the step of applying energy to theregion of weakness comprises irradiating the region of weakness with IRirradiation in an amount effective to induce fibrosis in a target layer.7. The method of claim 1, wherein the step of applying energy to theregion of weakness comprises irradiating the region of weakness withmicrowave irradiation in an amount effective to induce fibrosis in atarget layer.
 8. The method of claim 1, wherein the step of applyingenergy to the region of weakness comprises irradiating the region ofweakness with heat irradiation in an amount effective to induce fibrosisin a target layer.
 9. The method of claim 1, wherein the step ofapplying energy to the region of weakness comprises irradiating theregion of weakness with RF irradiation in an amount effective to inducefibrosis in a target layer.
 10. The method of claim 1, wherein themethod further comprises, administering a therapeutically effectiveamount of an agent to a subject, such that the agent is taken up by atleast one target layer of the vessel wall.
 11. The method of claim 10,wherein the method further comprises, administering a therapeuticallyeffective amount of photoactivatable agent, such that thephotoactivatable agent is activated upon irradiation to induce fibrosisin a target layer.
 12. A method of strengthening a vessel wall having anadventitial area in an area of an aneurysm of a subject comprising:administering a therapeutically effective amount of a photoactivatableagent to a subject, such that the agent is taken up by at least onelayer of the vessel wall of the aneurysm; applying energy to a targetregion of the vessel wall in an area of the aneurysm, such that thephotoactivatable agent is activated to strengthen the vessel wall; andincreasing an adventitial area of the vessel wall.
 13. The method ofclaim 12, wherein the step of administering a therapeutically effectiveamount of a photoactivatable agent further comprises systemicallyadministering the photoactivatable agent.
 14. The method of claim 12,wherein the step of administering a therapeutically effective amount ofa photoactivatable agent further comprises locally administering thephotoactivatable agent.
 15. The method of claim 12, wherein the step ofadministering a therapeutically effective amount of a photoactivatableagent further comprises administering a psoralen agent or derivativesthereof.
 16. The method of claim 12, wherein the step of applying energyto a target region further comprises irradiating the target regioninternally using a light delivery catheter.
 17. The method of claim 16,wherein the step of applying energy to a target region further comprisesirradiating the target region using a light delivery catheter withoutoccluding fluid flow.
 18. The method of claim 12, wherein the step ofapplying energy to a target region further comprises irradiating thetarget region externally using an external light delivery source. 19.The method of claim 12, wherein the step of applying energy to a targetregion further comprises irradiating the target region with UVirradiation.
 20. A method for increasing an adventitial area of a bloodvessel wall in an area of an aneurysm comprising: recognizing thatincreasing an adventitial area of a blood vessel strengthens the bloodvessel; administering a therapeutically effective amount of aphotoactivatable agent to a subject, such that the agent is taken up bythe adventitial area of the vessel in an area of the aneurysm; applyingenergy to a target region of the blood vessel wall so that thephotoactivatable agent is activated; and increasing the adventitial areaof the vessel in an area of the aneurysm.
 21. The method of claim 20,wherein the step of administering a therapeutically effective amount ofa photoactivatable agent further comprises systemically administeringthe photoactivatable agent.
 22. The method of claim 21, wherein the stepof administering a therapeutically effective amount of aphotoactivatable agent further comprises locally administering thephotoactivatable agent.
 23. The method of claim 21, wherein the step ofadministering a therapeutically effective amount of a photoactivatableagent further comprises administering a psoralen agent or derivativesthereof.
 24. The method of claim 21, wherein the step of applying energyto a target region further comprises irradiating the target regioninternally using a light delivery catheter.
 25. The method of claim 24,wherein the step of applying energy to a target region further comprisesirradiating the target region using a light delivery catheter withoutoccluding fluid flow.
 26. The method of claim 21, wherein the step ofapplying energy to a target region further comprises irradiating thetarget region externally using an external light delivery source. 27.The method of claim 21, wherein the step of applying energy to a targetregion further comprises irradiating the target region with UVirradiation.
 28. A method for treating an aneurysm by increasing anadventitial area of a blood vessel in an area of an aneurysm comprising:administering a therapeutically effective amount of a photoactivatableagent to a subject, such that the agent is taken up by an adventitialarea of the blood vessel in an area of the aneurysm; applying energy tothe site of the aneurysm to react within the photoactivatable agent; andincreasing an adventitial area in the area of the aneurysm.
 29. Themethod of claim 1, wherein the step of administering a therapeuticallyeffective amount of photoactivatable agent further comprisessystemically administering the photoactivatable agent.
 30. The method ofclaim 1, wherein the step of administering a therapeutically effectiveamount of photoactivatable agent further comprises locally administeringthe photoactivatable agent.
 31. The method of claim 1, wherein the stepof administering a therapeutically effective amount of photoactivatableagent further comprises administering a psoralen agent or derivativesthereof.
 32. The method of claim 30, wherein the step of applying energyto the site of the aneurysm further comprises irradiating the site ofthe aneurysm internally using a light delivery catheter.
 33. The methodof claim 32, wherein the step of applying energy to the site of theaneurysm further comprises irradiating the site of the aneurysminternally using a light delivery catheter without occluding fluid flow.34. The method of claim 30, wherein the step of applying energy to thesite of the aneurysm further comprises irradiating the site of theaneurysm externally using an external light delivery source.
 35. Themethod of claim 30, wherein the step of applying energy to the site ofthe aneurysm further comprises irradiating the site of the aneurysm withUV irradiation.
 36. A method for strengthening a vessel wall in an areaof an aneurysm of a subject comprising, irradiating a target region withUVC irradiation; and inducing fibrosis in at least one layer of thevessel wall in the area of the aneurysm.
 37. The method of claim 36,wherein the step of irradiating the target region further comprisesirradiating the target region internally using a light deliverycatheter.
 38. The method of claim 37, wherein the step of irradiatingthe target region further comprises irradiating the target regioninternally using a light delivery catheter without occluding fluid flow.39. The method of claim 36, wherein the step of irradiating the targetregion further comprises irradiating the target region externally usingan external light delivery source.
 40. The method of claim 36, whereinthe step of irradiating the target region further comprises irradiatingthe target region with UVC irradiation having a wavelength of about 240to 370 nanometers.
 41. A method for increasing an adventitial area of ablood vessel wall in an area of an aneurysm comprising, irradiating thetarget region with UVC irradiation; and increasing an adventitial areaof the blood vessel wall in the area of the aneurysm.
 42. The method ofclaim 41, wherein the step of irradiating the target region furthercomprises irradiating the target region internally using a lightdelivery catheter.
 43. The method of claim 42, wherein the step ofirradiating the target region further comprises irradiating the targetregion internally using a light delivery catheter without occludingfluid flow.
 44. The method of claim 41, wherein the step of irradiatingthe target region further comprises irradiating the target regionexternally using an external light delivery source.
 45. The method ofclaim 41, wherein the step of irradiating the target region furthercomprises irradiating the target region with UVC irradiation having awavelength of about 240 to 370 nanometers.
 46. A method for treating ananeurysm by increasing an adventitial area of a blood vessel in an areaof an aneurysm comprising, irradiating the site of the aneurysm with UVCirradiation; and increasing an adventitial area in the area of theaneurysm.
 47. The method of claim 46, wherein the step of irradiatingthe site of the aneurysm further comprises irradiating the site of theaneurysm internally using a light delivery catheter.
 48. The method ofclaim 47, wherein the step of irradiating the site of the aneurysmfurther comprises irradiating the site of the aneurysm internally usinga light delivery catheter without occluding the fluid flow.
 49. Themethod of claim 46, wherein the step of irradiating the site of theaneurysm further comprises irradiating the site of the aneurysmexternally using an external light delivery source.
 50. The method ofclaim 46, wherein the step of irradiating the site of the aneurysmfurther comprises irradiating the site of the aneurysm with UVCirradiation having a wavelength of about 240 to 370 nanometers.